Population Imaging of Action Potentials by Novel Two-Photon Microscopes and Genetically Encoded Voltage Indicators

通过新型双光子显微镜和基因编码电压指示器对动作电位进行群体成像

• 批准号: 9588470
• 负责人: Jerry L Chen
• 金额: $ 268.09万
• 依托单位: BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2022-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9588470
• 关键词: Action Potentials; Address; Anatomy; Animals; Behavior; Biological; Brain; Brain Diseases; Calcium; Cells; Code; Communication; Communities; Development; Electrophysiology (science); Elements; Engineering; Fiber; Frequencies; Future; Genetic Engineering; Goals; Image; Individual; Interneurons; Investigation; Kinetics; Label; Lasers; Measurement; Measures; Membrane; Methods; Microscope; Microscopy; Molecular; Monitor; Mus; Neocortex; Neurons; Neurosciences; Noise; Opsin; Optical Methods; Optics; Phase; Photons; Physiologic pulse; Population; Process; Property; Protein Engineering; Reporting; Research Personnel; Resolution; Scanning; Signal Transduction; Source; Speed; System; Techniques; Technology; Testing; Thulium; Time; Training; Vibrissae; Work; awake; base; design; detector; experimental study; genetic information; imaging system; in vivo; in vivo evaluation; instrumentation; millisecond; neuronal circuitry; neuronal patterning; new technology; non-invasive imaging; novel; operation; optical imaging; relating to nervous system; sapphire laser; scale up; screening; sensor; somatosensory; tool; two-photon; voltage

PROJECT SUMMARY Understanding how information is processed in the mammalian neocortex has been a longstanding question in neuroscience. While the action potential is the fundamental bit of information, how these spikes encode representations and drive behavior remains unclear. In order to adequately address this problem, it has become apparent that experiments are needed in which activity from large numbers of neurons can be measured in a detailed and comprehensive manner across multiple timescales. Direct measurements of action potentials have primarily been achieved by electrophysiology. However, such measurements cannot easily be combined with other methods to assess the connectivity and molecular properties of neurons. Integrating functional, anatomical, and genetic information is critical for understanding how neuronal circuits are organized and computed. There have been long-standing efforts in developing optical methods for measuring neuronal activity due to its compatibility to simultaneously measure connectivity and molecular identity using fluorescent labeling techniques. Newly engineered genetically-encoded voltage-sensitive indicators have now opened the door for optical imaging of action potentials. Two-photon microscopy has been a proven method for deep non-invasive imaging into the brain. However, the fast millisecond transience of action potentials and the membrane localization of genetically-encoded voltage-sensitive indicators both contribute to conditions of limited photon flux. This creates fundamental challenges in the application of two- photon microscopy for voltage imaging that requires scanning at kilohertz frame rates with high signal to noise. To achieve this requires a concerted effort between optical engineers and protein engineers to develop new instrumentation and sensors to arrive at an optimal solution. This multi-investigator effort proposes to advance two-photon microscopy and genetically-encoded voltage-sensitive indicators to enable non-invasive population-level measurements of action potentials with single-cell spatial resolution and single-spike temporal precision deep into the mammalian brain of awake behaving animals.

项目摘要 了解在哺乳动物新皮层中如何处理信息是一个长期存在的问题 神经科学。虽然动作电位是基本信息，但这些尖峰如何编码 表示和驱动行为尚不清楚。为了充分解决这个问题， 显然需要实验，其中大量神经元的活动可能是 在多个时间尺度上以详细而全面的方式进行测量。直接测量 动作电位主要是通过电生理学实现的。但是，这样的测量不能 很容易与其他方法结合使用，以评估神经元的连通性和分子特性。 整合功能，解剖学和遗传信息对于理解神经元电路至关重要 是组织和计算的。在开发光学方法方面已经做出了长期的努力 由于其同时测量连通性和分子的兼容性，测量神经元活性 使用荧光标签技术的身份。新设计的遗传编码的电压敏感 现在，指标已经为动作电位的光学成像打开了大门。两光子显微镜具有 是对大脑进行深度非侵入成像的验证方法。但是，快速的毫秒短暂性 作用电位和遗传编码电压敏感指标的膜定位都 有助于光子通量有限的条件。这在应用两个方面构成了基本挑战 用于电压成像的光子显微镜需要以高信号到噪声的kilohertz帧速率进行扫描。 为了实现这一目标，需要光学工程师和蛋白质工程师之间的一致努力来开发新的 仪器和传感器可得出最佳解决方案。这项多次评估者的努力提出了进步 两光子显微镜和遗传编码的电压敏感指标，使得无创的 通过单细胞空间分辨率和单峰时间的人群级测量的动作电位测量 精确地深入到醒着的动物的哺乳动物大脑。